The following description is provided to assist the understanding of the reader. None of the information provided or references cited is admitted to be prior art to the present invention.
Genetic abnormalities e.g. duplication, deletion, chromosomal translocation, point mutation often leads to pathological conditions.
Some diseases, such as cancer, are due to genetic abnormalities acquired in a few cells during life, while in other diseases the genetic abnormality is present in all cells of the body and present since conception.
To detect a genetic abnormality it is necessary to detect the genomic nucleic acid containing the abnormality. Methods of detection of genetic abnormalities, such as aneuploidy, translocations, duplications and deletions are well known in the art. One such methods includes cytogenetic analysis in which a metaphase spread of chromosomes is stained and visualized. Metaphase chromosomes exhibit a particular pattern of light and dark staining manifested in a chromosomal banding pattern.
The development of molecular cytogenetic approaches offer assays with greater sensitivity. These techniques incorporate DNA hybridization with radiolabeled or fluorescent labeled probes. For example, in fluorescence in situ hybridization (FISH) analysis, a fluorescent probe is hybridized to metaphase or interphase chromosomes. The hybridized probe is then detected using a fluorescence microscope. However, these methods require intact cells or intact or partially intact nuclei.
Non-in situ hybridization methods of detecting chromosomal abnormalities are also known in the art. US Patent publication 2006/0292576 describes a method of detecting chromosomal abnormalities by hybridizing, capturing, and detecting genomic DNA on a solid support. This method utilizes two probes specific for the genomic nucleic acid. One probe is anchored to the solid support and the other probe is detectably labeled. Hybridization of the first probe with the genomic nucleic acid captures the genomic nucleic acid on the solid support, while the signal from the detectable labels of the second probe is used to detect the genomic nucleic acid. This method require capturing of genomic nucleic acid to solid support by nucleic acid hybridization.
One method of detecting a single nucleotide polymorphism is by dynamic allele-specific hybridization (DASH). This method takes advantage of the differences in the melting temperature (Tm) in DNA that results from the instability of mismatched base pairs. The genomic segment is amplified e.g. by PCR and attached to a solid support. An allele specific oligonucleotide is added in the presence of a molecule that fluoresces when bound to double-stranded DNA. The intensity is measured as temperature is increased until the Tm is determined. A mutation will result in a lower than expected Tm and thus can be distinguished from the wild type sequence.
Another method of detecting a single nucleotide polymorphism is by microarray technology. The genomic nucleic acid is initially amplified, then captured on a solid surface by hybridization to a probe anchored to the solid surface. The genomic nucleic acid is detected by hybridization using a second probe with a detectable label.